3,139 research outputs found
Ozonolysis of surface-adsorbed methoxyphenols: kinetics of aromatic ring cleavage vs. alkene side-chain oxidation
Lignin pyrolysis products, which include a variety of substituted
methoxyphenols, constitute a major component of organics released by biomass
combustion, and may play a central role in the formation of atmospheric brown
carbon. Understanding the atmospheric fate of these compounds upon exposure
to trace gases is therefore critical to predicting the chemical and physical
properties of biomass burning aerosol. We used diffuse reflectance infrared
spectroscopy to monitor the heterogeneous ozonolysis of 4-propylguaiacol,
eugenol, and isoeugenol adsorbed on NaCl and α-Al<sub>2</sub>O<sub>3</sub> substrates.
Adsorption of gaseous methoxyphenols onto these substrates produced near-monolayer surface concentrations of
3 × 10<sup>18</sup> molecules m<sup>−2</sup>. The subsequent dark
heterogeneous ozonolysis of adsorbed 4-propylguaiacol cleaved the aromatic
ring between the methoxy and phenol groups with the product conclusively
identified by GC-MS and <sup>1</sup>H-NMR. Kinetic analysis of eugenol and
isoeugenol dark ozonolysis also suggested the formation of ring-cleaved
products, although ozonolysis of the unsaturated substituent groups forming
carboxylic acids and aldehydes was an order of magnitude faster. Average
uptake coefficients for NaCl-adsorbed methoxyphenols were
γ = 2.3 (± 0.8) × 10<sup>−7</sup> and
2 (± 1) × 10<sup>−6</sup> for ozonolysis of the aromatic ring and
the unsaturated side chain, respectively, and reactions on
α-Al<sub>2</sub>O<sub>3</sub> were approximately two times slower. UV–visible
radiation (λ > 300 nm) enhanced eugenol ozonolysis of the
aromatic ring by a factor of 4(± 1) but had no effect on ozonolysis of
the alkene side chain
Effect of cyclosporine on hepatic cytosolic estrogen and androgen receptor levels before and after partial hepatectomy
Estrogen and androgen receptors within the liver have been reported to modulate the hepatic regenerative response to partial hepatectomy. Moreover, cyclosporine has several untoward effects that might occur as a consequence of alterations in sex hormone activity. To evaluate these questions the following experiments were performed. Estrogen and androgen receptors in cytosol were quantitated in livers of rats treated with cyclosporine or olive oil vehicle before and after partial hepatectomy or a sham operation. Ornithine decarboxylase activity and thymidine kinase activity were assessed as indices of hepatic regeneration. Preoperative levels of estrogen receptor activity in the hepatic cytosol were significantly greater in rats treated with cyclosporine as compared to vehicle treated controls (P<0.01). In contrast, preoperative levels of androgen receptor activity in the cyclosporine-treated and vehicle-treated animals were similar. Following partial hepatectomy, a reduction in the activity of both sex hormone receptors in the hepatic cytosol was observed and was compatible with results described previously in normal animals. Unexpectedly the preoperative levels of ornithine decarboxylase (P<0.01) and thymidine kinase activity (P<0.01) were significantly greater in the rats treated with cyclosporine as compared to the vehicle treated controls. As expected, ornithine decarboxylase activity (at 6 hr) and thymidine kinase activity (at 24 hr) rose and peaked in response to a partial hepatectomy but were significantly greater (P<0.05) in the rats treated with cyclosporine as compared to the vehicle. These results show that cyclosporine treatment causes an increase in the hepatic content of estrogen receptor activity that is associated with an enhanced potential for a regenerative response. These effects of cyclosporine treatment on the sex hormone receptor levels in liver may explain the mechanisms responsible for some of the untoward effects of treatment with this agent. © 1990 Plenum Publishing Corporation
Utility of the first few100 approach during the 2009 influenza A(H1N1) pandemic in the Netherlands
Microbially mediated reduction of FeIII and AsV in Cambodian sediments amended with 13C-labelled hexadecane and kerogen
Microbial activity is generally accepted to play a critical role, with the aid of suitable organic carbon substrates, in the mobilisation of arsenic from sediments into shallow reducing groundwaters. The nature of the organic matter in natural aquifers driving the reduction of AsV to AsIII is of particular importance but is poorly understood. In this study, sediments from an arsenic rich aquifer in Cambodia were amended with two 13C-labelled organic substrates. 13C-hexadecane was used as a model for potentially bioavailable long chain n-alkanes and a 13C-kerogen analogue as a proxy for non-extractable organic matter. During anaerobic incubation for 8 weeks, significant FeIII reduction and AsIII mobilisation were observed in the biotic microcosms only, suggesting that these processes were microbially driven. Microcosms amended with 13C-hexadecane exhibited a similar extent of FeIII reduction to the non-amended microcosms, but marginally higher AsIII release. Moreover, gas chromatography–mass spectrometry analysis showed that 65 % of the added 13C-hexadecane was degraded during the 8-week incubation. The degradation of 13C-hexadecane was microbially driven, as confirmed by DNA stable isotope probing (DNA-SIP). Amendment with 13C-kerogen did not enhance FeIII reduction or AsIII mobilisation, and microbial degradation of kerogen could not be confirmed conclusively by DNA-SIP fractionation or 13C incorporation in the phospholipid fatty acids. These data are, therefore, consistent with the utilisation of long chain n-alkanes (but not kerogen) as electron donors for anaerobic processes, potentially including FeIII and AsV reduction in the subsurface
Some Secrets of Fluorescent Proteins: Distinct Bleaching in Various Mounting Fluids and Photoactivation of cyan fluorescent proteins at YFP-Excitation
Background
The use of spectrally distinct variants of green fluorescent protein (GFP) such as cyan or yellow mutants (CFP and YFP, respectively) is very common in all different fields of life sciences, e.g. for marking specific proteins or cells or to determine protein interactions. In the latter case, the quantum physical phenomenon of fluorescence resonance energy transfer (FRET) is exploited by specific microscopy techniques to visualize proximity of proteins.

Methodology/Principal Findings
When we applied a commonly used FRET microscopy technique - the increase in donor (CFP)-fluorescence after bleaching of acceptor fluorophores (YFP), we obtained good signals in live cells, but very weak signals for the same samples after fixation and mounting in commercial microscopy mounting fluids. This observation could be traced back to much faster bleaching of CFP in these mounting media. Strikingly, the opposite effect of the mounting fluid was observed for YFP and also for other proteins such as Cerulean, TFP or Venus. The changes in photostability of CFP and YFP were not caused by the fixation but directly dependent on the mounting fluid. Furthermore we made the interesting observation that the CFP-fluorescence intensity increases by about 10 - 15% after illumination at the YFP-excitation wavelength – a phenomenon, which was also observed for Cerulean. This photoactivation of cyan fluorescent proteins at the YFP-excitation can cause false-positive signals in the FRET-microscopy technique that is based on bleaching of a yellow FRET acceptor.

Conclusions/Significance
Our results show that photostability of fluorescent proteins differs significantly for various media and that CFP bleaches significantly faster in commercial mounting fluids, while the opposite is observed for YFP and some other proteins. Moreover, we show that the FRET microscopy technique that is based on bleaching of the YFP is prone to artifacts due to photoactivation of cyan fluorescent proteins under these conditions
Endothelial repair in stented arteries is accelerated by inhibition of Rho-associated protein kinase.
AIMS: Stent deployment causes endothelial cell (EC) denudation, which promotes in-stent restenosis and thrombosis. Thus endothelial regrowth in stented arteries is an important therapeutic goal. Stent struts modify local hemodynamics, however the effects of flow pertubation on EC injury and repair are incompletely understood. By studying the effects of stent struts on flow and EC migration we identified an intervention that promotes endothelial repair in stented arteries. METHODS AND RESULTS: In vitro and in vivo models were developed to monitor endothelialization under flow and the influence of stent struts. A 2D parallel-plate flow chamber with 100 μm ridges arranged perpendicular to the flow was used. Live cell imaging coupled to computational fluid dynamic simulations revealed that EC migrate in the direction of flow upstream from the ridges but subsequently accumulate downstream from ridges at sites of bidirectional flow. The mechanism of EC trapping by bidirectional flow involved reduced migratory polarity associated with altered actin dynamics. Inhibition of Rho-associated protein kinase (ROCK) enhanced endothelialization of ridged surfaces by promoting migratory polarity under bidirectional flow (p<0.01). To more closely mimic the in vivo situation we cultured EC on the inner surface of polydimethylsiloxane tubing containing Coroflex Blue stents (65 μm struts) and monitored migration. ROCK inhibition significantly enhanced EC accumulation downstream from struts under flow (p<0.05). We investigated the effects of ROCK inhibition on re-endothelialization in vivo using a porcine model of EC denudation and stent placement. En face staining and confocal microscopy revealed that inhibition of ROCK using fasudil (30 mg/day via osmotic minipump) significantly increased re-endothelialization of stented carotid arteries (p<0.05). CONCLUSIONS: Stent struts delay endothelial repair by generating localised bidirectional flow which traps migrating EC. ROCK inhibitors accelerate endothelial repair of stented arteries by enhancing EC polarity and migration through regions of bidirectional flow
Cytosine-to-Uracil Deamination by SssI DNA Methyltransferase
The prokaryotic DNA(cytosine-5)methyltransferase M.SssI shares the specificity of eukaryotic DNA methyltransferases (CG) and is an important model and experimental tool in the study of eukaryotic DNA methylation. Previously, M.SssI was shown to be able to catalyze deamination of the target cytosine to uracil if the methyl donor S-adenosyl-methionine (SAM) was missing from the reaction. To test whether this side-activity of the enzyme can be used to distinguish between unmethylated and C5-methylated cytosines in CG dinucleotides, we re-investigated, using a sensitive genetic reversion assay, the cytosine deaminase activity of M.SssI. Confirming previous results we showed that M.SssI can deaminate cytosine to uracil in a slow reaction in the absence of SAM and that the rate of this reaction can be increased by the SAM analogue 5’-amino-5’-deoxyadenosine. We could not detect M.SssI-catalyzed deamination of C5-methylcytosine (m5C). We found conditions where the rate of M.SssI mediated C-to-U deamination was at least 100-fold higher than the rate of m5C-to-T conversion. Although this difference in reactivities suggests that the enzyme could be used to identify C5-methylated cytosines in the epigenetically important CG dinucleotides, the rate of M.SssI mediated cytosine deamination is too low to become an enzymatic alternative to the bisulfite reaction. Amino acid replacements in the presumed SAM binding pocket of M.SssI (F17S and G19D) resulted in greatly reduced methyltransferase activity. The G19D variant showed cytosine deaminase activity in E. coli, at physiological SAM concentrations. Interestingly, the C-to-U deaminase activity was also detectable in an E. coli ung+ host proficient in uracil excision repair
Impaired neutralising antibody formation and high transduction efficacy after isolated hepatic perfusion with adenoviral vectors
Local adenoviral gene transfer can be performed by means of isolated hepatic perfusion (IHP). This methodology is a very effective and safe way to deliver adenoviral vectors. We studied the immune response after IHP, A decreased neutralising antibody formation was observed, offering possibilities for further research in the field of gene therapy in isolated perfusion settings
Phenotypic Variation and Bistable Switching in Bacteria
Microbial research generally focuses on clonal populations. However, bacterial cells with identical genotypes frequently display different phenotypes under identical conditions. This microbial cell individuality is receiving increasing attention in the literature because of its impact on cellular differentiation, survival under selective conditions, and the interaction of pathogens with their hosts. It is becoming clear that stochasticity in gene expression in conjunction with the architecture of the gene network that underlies the cellular processes can generate phenotypic variation. An important regulatory mechanism is the so-called positive feedback, in which a system reinforces its own response, for instance by stimulating the production of an activator. Bistability is an interesting and relevant phenomenon, in which two distinct subpopulations of cells showing discrete levels of gene expression coexist in a single culture. In this chapter, we address techniques and approaches used to establish phenotypic variation, and relate three well-characterized examples of bistability to the molecular mechanisms that govern these processes, with a focus on positive feedback.
Experimental Demonstration of the Fitness Consequences of an Introduced Parasite of Darwin's Finches
Introduced parasites are a particular threat to small populations of hosts living on islands because extinction can occur before hosts have a chance to evolve effective defenses. An experimental approach in which parasite abundance is manipulated in the field can be the most informative means of assessing a parasite's impact on the host. The parasitic fly Philornis downsi, recently introduced to the Galápagos Islands, feeds on nestling Darwin's finches and other land birds. Several correlational studies, and one experimental study of mixed species over several years, reported that the flies reduce host fitness. Here we report the results of a larger scale experimental study of a single species at a single site over a single breeding season.We manipulated the abundance of flies in the nests of medium ground finches (Geospiza fortis) and quantified the impact of the parasites on nestling growth and fledging success. We used nylon nest liners to reduce the number of parasites in 24 nests, leaving another 24 nests as controls. A significant reduction in mean parasite abundance led to a significant increase in the number of nests that successfully fledged young. Nestlings in parasite-reduced nests also tended to be larger prior to fledging.Our results confirm that P. downsi has significant negative effects on the fitness of medium ground finches, and they may pose a serious threat to other species of Darwin's finches. These data can help in the design of management plans for controlling P. downsi in Darwin's finch breeding populations
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